Functional studies of protease inhibitors using tick and mosquito cell cultures

Abstract

The present propose is included in the thematic project with the following aim: study of the physiological role of proteases and their inhibitors identified in ectoparasites that concern public health and livestock production. Among these vectors, we have been studied: the tick Rhipicephalus (Boophilus) microplus, responsible for the transmission of Babesia bovis protozoa and Anaplasma marginale bacteria for bovines; and the mosquito Aedes aegypti, vector of Dengue, Zika and Chikungunya virus. Shredding light in the biological functions of molecules that take part in host-parasite biology may help the development of new drugs and pesticides for vector control. Thus, we have selected two molecules for further studies. The first one is a cisteine protease inhibitor from A. aegypti, which was found to be modulated during DENV2 infection, and the latter a molecule found in the saliva of R. microplus, named BmSEI (Boophilus microplus Saliva Elastase Inhibitor), which is similar to antimicrobial peptides. Recent experiments reveal the presence of BmSEI transcripts in several tick tissues such as midgut, ovary and fat body. The first objective of this project is the establishment of BME26 and Aag2 cell cultures from R. microplus and A. aegypti, respectively, in our lab, located in the Departamento de Bioquímica da Escola Paulista de Medicina - UNIFESP. This project phase will be performed in collaboration with Dra. Sirlei Daffre of Departamento de Imunologia - ICB - USP and Dr. Marcos Sorgine of Instituto de Bioquímica Médica - UFRJ. To assess BmSEI function, RNAi in BME26 culture will be performed, followed by transcriptome analysis of silenced and non-silenced cells. Likewise, RNAi of A. aegypti cysteine protease inhibitor will be carried in Aag2 cell line and its relation with apoptose pathway analyzed. (AU)